Fig. 3
GABA-containing ADNVs suppress the inflammatory signaling. (A, B) GC-MS analysis of GABA in ADNVs according to the reference substance (A), and the concentrations were calculated. (C) qPCR analysis of the indicated cytokines in MH-S cells that were pre-incubated with ADNVs (GABA equal to 4 ng/mL) or GABA (100 nM), followed by LPS (100 ng/mL) stimulation for 4 h. (D, E) Western-blotting of the indicated signaling molecules in MH-S cells that were pre-incubated with ADNVs (GABA equal to 4 ng/mL) or GABA (100 nM) and then stimulated with LPS (100 ng/mL) for 4 h.(F, G)Western-blotting of the indicated signaling molecules in MH-S cells as described in D. Alternatively, bicuculline (Bic, 30 µM) or saclofe (Sac, 15 µM) were additionally added to the cellular culture. (H) The intracellular Ca2+ levels were measured as described in Methods. (I) Western-blotting of pGSK-3β/GSK-3β in MH-S cells as described in D. Bicuculline (Bic, 30 µM) or saclofe (Sac, 15 µM) were added in some circumstances. Molecular weights of the indicated molecules are depicted. The results are one of three independent experiments. Shown are representative images, and the data are expressed as the mean ± SD, *P < 0.05, **P < 0.01, ***P < 0.001